Uridine insertion/deletion RNA editing is an extensive post-transcriptional modification of mitochondrial mRNAs in kinetoplastid organisms, including Trypanosoma brucei This process is carried out using trans-acting gRNAs and complex protein machinery. The essential RNA editing substrate binding complex (RESC) serves as the scaffold that modulates protein and RNA interactions during editing, and contains the guide RNA binding complex (GRBC), the RNA editing mediator complexes (REMCs), and organizer proteins. Despite the importance of RESC in editing, the functions of each protein comprising this complex are not completely understood. Here, we further define the roles of a REMC protein, RESC13, and a RESC organizer, RESC14, using high-throughput sequencing on two large pan-edited mRNAs, A6 and COIII. When comparing our analyses to that of a previously published small pan-edited mRNA, RPS12, we find that RESC13 has conserved functions across the three transcripts with regard to editing initiation, gRNA utilization, gRNA exchange, and restricting the formation of long misedited junctions that likely arise from its ability to modulate RNA structure. However, RESC13 does have transcript-specific effects on the types of long junctions whose formation it restricts. RESC14 has a conserved effect on gRNA utilization across the three transcripts analyzed, but has transcript-specific effects on editing initiation, gRNA exchange, and junction formation. Our data suggest that transcript-specific effects of both proteins are due to differences in transcript length and sequences as well as transcript-specific protein interactions. These findings highlight the importance of studying multiple transcripts to determine the function of editing factors.
Keywords: Trypanosoma brucei; U-indel RNA editing; high-throughput sequencing; holoenzyme; trypanosome.
© 2022 Sortino et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.